Development of multivalent, ultrapotent nanobody cocktails for SARS-CoV-2 neutralization

开发用于中和 SARS-CoV-2 的多价、超强纳米抗体混合物

• 批准号: 10444442
• 负责人: William Paul Duprex
• 金额: $ 76.9万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-02-09 至 2022-02-18
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10444442
• 关键词: 2019-nCoV; Aerosols; Affinity; Aftercare; Antibodies; Binding; Biological Availability; Biomedical Engineering; COVID-19; COVID-19 outbreak; COVID-19 therapeutics; COVID-19 treatment; Cells; Clinical; Clinical Research; Clinical Trials; Communities; Computer software; Coronavirus; Deposition; Development; Diagnostic Reagent; Dose; Drug Storage; Epitope Mapping; Epitopes; Event; Future; Genotype; Goals; Hamsters; Human; Immunoglobulin Fragments; Immunoglobulin G; Infection; Inhalation; Innovative Therapy; Lung; Microbe; Modeling; Mutation; Nebulizer; Pharmaceutical Preparations; Primates; Production; Property; Proteomics; Pulmonary Pathology; Reporting; Research; Resistance; Resolution; Respiratory System; SARS-CoV-2 infection; SARS-CoV-2 variant; Science; Structure; System; Technology; Therapeutic; Therapeutic Agents; Translations; Viral; Viral Antibodies; Viral Load result; Viral Pneumonia; Virus; aerosolized; base; clinical application; cost; cost effective; design; drug development; drug quality; efficacy evaluation; emerging pathogen; flexibility; global health; improved; in vivo; insight; interdisciplinary approach; interest; multidisciplinary; nanobodies; novel; novel therapeutics; novel vaccines; pandemic disease; pathogen; preclinical efficacy; preclinical evaluation; prevent; protein folding; rational design; receptor binding; respiratory virus; structural biology; therapeutic candidate; therapeutically effective; translational potential; variants of concern

PROJECT SUMMARY/ABSTRACT The outbreak of COVID-19 has severely impacted global health and the economy. Cost-effective, highly efficacious therapeutics are urgently needed. Camelid VHH antibodies or nanobodies (Nbs) are small, highly stable, easily bioengineered, and can be rapidly and economically manufactured from microbes. They are highly robust and are flexible for administration, including possible delivery by nebulization. Together these unique properties of Nbs make their uses against respiratory viruses such as SARS-CoV-2 especially appealing. We recently developed a disruptive proteomic technology for large-scale identification of multi-epitope, drug- quality Nbs (Xiang et. al, Cell Systems. 2021). Using this technology, we identified > 8,000 high-affinity Nbs for the SARS-CoV-2 spike (S) receptor-binding domain (RBD) including Nbs that target highly neutralizing epitopes with sub-pM affinities and can neutralize SARS-CoV-2 at sub-ng/ml concentrations, which are unprecedented for antiviral antibody fragments. Structural proteomics revealed multiple distinct epitopes and potential neutralization mechanisms. Bioengineering of multi-epitope and multivalent constructs improved the potency to below 0.1 ng/ml (Xiang, et. al, Science. 2020). Most recently, we have demonstrated the high preclinical efficacy of an ultrapotent and stable trimeric Nb construct (PiN-21) for inhalation treatment of SARS- CoV-2 infection in a sensitive COVID-19 model (Nambulli, et. al, Science Advances. 2021). Intranasal delivery of PiN-21 at 0.6 mg/kg substantially reduces viral burdens in both airways. Critically, aerosol delivery of PiN-21 at 0.2 mg/kg decreases lung viral titers by 6-logs, minimizing lung pathology post-infection and preventing viral pneumonia. Combined with the marked stability and low production cost, this innovative therapy may provide a convenient and cost-effective option to mitigate the evolving pandemic and future events. In the revision, we aim to identify and characterize highly potent Nbs that are highly resistant to the variants of concern (VOCs) of SARS-CoV-2, investigate the neutralization mechanisms by structural approaches, and develop ultrapotent Nb constructs into safe and effective therapeutics. Our central hypothesis is that Nbs can be bioengineered into multivalent and ultrapotent forms to resist the mutational escape and the variants of concerns (VOCs) of SARS-CoV-2. Completion of our proposed studies will lead to cost-effective and convenient COVID-19 therapeutic candidates for translation into clinical trials. High-resolution structural studies will provide critical insights into how Nbs uniquely target the virus for high-affinity binding and neutralization. Critically, this project will serve as the testbed of our multidisciplinary platform to develop potent therapeutic and diagnostic reagents for future pandemics caused by coronaviruses or other pathogens.

项目概要/摘要 COVID-19 的爆发严重影响了全球健康和经济。性价比高， 迫切需要有效的治疗方法。骆驼科动物 VHH 抗体或纳米抗体 (Nbs) 体积小、高度 稳定、易于生物工程，并且可以通过微生物快速、经济地制造。他们是 高度稳健且给药灵活，包括可能通过雾化给药。一起这些 Nbs 的独特特性使其可用于对抗呼吸道病毒，例如 SARS-CoV-2 有吸引力的。 我们最近开发了一种破坏性蛋白质组技术，用于大规模鉴定多表位、药物 质量 Nbs（Xiang 等人，Cell Systems.2021）。利用这项技术，我们鉴定了超过 8,000 个高亲和力 Nb SARS-CoV-2 刺突 (S) 受体结合域 (RBD)，包括针对高度中和的 Nb 具有亚 pM 亲和力的表位，可以在亚 ng/ml 浓度下中和 SARS-CoV-2，其中 抗病毒抗体片段前所未有。结构蛋白质组学揭示了多个不同的表位和 潜在的中和机制。多表位和多价构建体的生物工程改善了 效力低于 0.1 ng/ml（Xiang 等人，Science.2020）。最近，我们展示了高 超强稳定三聚 Nb 构建体 (PiN-21) 用于吸入治疗 SARS 的临床前疗效 敏感的 COVID-19 模型中的 CoV-2 感染（Nambulli 等人，Science Advances.2021）。鼻内给药 0.6 mg/kg 的 PiN-21 显着降低了两个气道中的病毒负荷。至关重要的是，PiN-21 的气雾输送 0.2 mg/kg 可使肺部病毒滴度降低 6 个对数级，最大限度地减少感染后的肺部病理并预防病毒 肺炎。结合显着的稳定性和较低的生产成本，这种创新疗法可能会提供一种 缓解不断变化的流行病和未来事件的便捷且具有成本效益的选择。 在修订中，我们的目标是识别和表征对以下变异体具有高度抵抗力的高效 Nbs。 SARS-CoV-2 的关注点（VOC），通过结构方法研究中和机制，以及 将超强铌结构开发成安全有效的疗法。我们的中心假设是 Nbs 可以 被生物工程改造为多价和超强形式，以抵抗突变逃逸和变异 SARS-CoV-2 的关注点（VOC）。 完成我们提出的研究将带来具有成本效益且方便的 COVID-19 治疗方法 转化为临床试验的候选者。高分辨率结构研究将为以下问题提供重要见解 Nbs 如何独特地针对病毒进行高亲和力结合和中和。至关重要的是，该项目将作为 我们的多学科平台的试验台，用于开发未来有效的治疗和诊断试剂 由冠状病毒或其他病原体引起的流行病。